ABSTRACT Systemic Sclerosis (SSc) is a systemic autoimmune disease of unknown origin characterized by progressive fibrosis affecting skin and internal organs leading to severe disability, multiple organ failure, and a high mortality rate. Pulmonary involvement is the leading cause of functional disability and high mortality in SSc. A major unmet need for SSc clinical management is the absence of well validated biomarkers that allow early diagnosis and accurate assessment of SSc severity and extent of involvement, or that differentiate patients with Primary Raynaud Phenomenon from patients with Raynaud Phenomenon at risk of evolving into SSc. We recently obtained strong Preliminary Results indicating that microRNA (miRNA) analysis of exosomes isolated from the serum of SSc patients may provide unique and sensitive biomarkers that reflect the extent and severity of SSc. Based on these results we propose the following hypothesis: ?The unbiased analysis of the miRNA profile of exosomes isolated from SSc patient serum will allow to separate with a high degree of accuracy and sensitivity various relevant clinical subsets of SSc, will display a significant correlation with SSc severity and extent of pulmonary fibrotic involvement, and will distinguish patients with Primary Raynaud Phenomenon from patients with Raynaud Phenomenon harboring serum antinuclear antibodies (ANA) who are at high risk of evolving into SSc. A further hypothesis posits that: ?The differentially expressed miRNA identified in SSc serum exosomes play a crucial role in the pathogenesis of SSc tissue fibrosis inducing profibrotic molecular events on normal fibroblasts or inhibiting the profibrotic phenotype of SSc fibroblasts.? To test these hypotheses we will pursue the following Specific Aims: SPECIFIC AIM 1: To identify employing Next Generation Sequencing (NGS) differences in the miRNA profile of exosomes isolated from the serum of the following individuals: A: Patients with recent onset diffuse SSc vs. normal individuals; B: Patients with SSc-associated pulmonary fibrosis vs. diffuse SSc patients without lung involvement; and C: Patients with Primary Raynaud Phenomenon vs. patients with ANA-positive Raynaud Phenomenon. SPECIFIC AIM 2: To examine whether the most differentially expressed (upregulated or downregulated) miRNA identified in Aim 1 exert either profibrotic effects on normal dermal and lung fibroblasts or inhibit the fibrotic phenotype or SSc dermal and lung fibroblasts in vitro. The unbiased NGS miRNA analysis of serum exosomes proposed here is entirely novel and has not been applied to SSc previously. The differences in miRNAs present in serum exosomes identified here may lead to the future development of non-invasive SSc biomarkers that will result in a ?transformational change? in the standard of care for SSc patients and for patients with Raynaud Phenomenon at high risk of evolving into SSc. We further expect that the mechanistic studies performed with the most differentially expressed serum exosome miRNAs will identify novel molecular mechanisms and pathways involved in SSc tissue fibrosis.